AA–LTB4–ROS Axis drives macrophage extracellular trap formation and endothelial injury in smoking-induced pulmonary hypertension
Y Wu, Q ZhouAbstract
Background
Smoking-associated pulmonary hypertension (PH) is characterized by chronic pulmonary vascular inflammation, endothelial dysfunction[1], and vascular remodeling. Growing evidence suggests that macrophage dysregulation and lipid metabolic signaling, particularly arachidonic acid (AA)–derived leukotriene B4 (LTB4), play a critical role in driving endothelial injury[2]. Despite this, the mechanistic link between cigarette smoke exposure, macrophage extracellular trap (MET) formation, and subsequent endothelial dysfunction remains poorly defined.
Purpose
We aimed to determine whether cigarette smoke triggers AA metabolic remodeling in macrophages, leading to LTB4-dependent reactive oxygen species (ROS) generation via NOX2, PAD2-mediated MET formation, and downstream endothelial apoptosis, barrier disruption, and inflammatory activation in smoking-related PH.
Methods
Single-cell transcriptomic datasets and lung tissues from PH patients were analyzed for macrophage abundance, AA metabolic gene expression, and MET deposition. A chronic cigarette smoke (CS)-exposed mouse model was used to evaluate pulmonary vascular remodeling, right ventricular function, and hemodynamic parameters. CS-stimulated macrophages underwent untargeted metabolomic profiling, ROS measurement, and MET formation assays. Macrophage–endothelial co-cultures were used to assess endothelial apoptosis, barrier integrity, and inflammatory mediator expression. Pharmacological inhibitors targeting AA metabolism, NOX2/ROS, or PAD2 were applied to probe pathway dependency.
Results
PH lungs exhibited marked macrophage enrichment, upregulated AA metabolic genes, and enhanced macrophage–endothelial interactions. Lung tissues from smoking PH patients showed elevated MET deposition, evidenced by citrullinated histone H3, myeloperoxidase, and extracellular DNA. CS-exposed mice developed pulmonary arterial thickening, perivascular collagen accumulation, reduced PAT/PET ratio, decreased TAPSE, and right ventricular hypertrophy, consistent with pulmonary vascular remodeling and right heart dysfunction. In vitro, CSE enhanced macrophage LTB4 production, NOX2-mediated ROS, and PAD2-driven MET formation, leading to endothelial apoptosis, barrier compromise, and increased inflammatory signaling. Inhibition of AA metabolism, ROS, or PAD2 significantly mitigated MET formation and preserved endothelial function.
Conclusion
Cigarette smoke induces AA metabolic remodeling in macrophages, activating an LTB4–NOX2–ROS–PAD2–MET axis that promotes endothelial injury and pulmonary vascular remodeling. Targeting macrophage lipid metabolism and MET formation may provide a promising therapeutic avenue for smoking-associated PH.